1. Field of the Invention
This invention relates to the field of diagnosis and therapy of cancer and the prevention and treatment of viral infections. More particularly, it relates to a polypeptide having the antibody binding specificity of the 46 Kdalton HMFG antigen, hybrid protein thereof, anti-idiotype antibodies and polynucleotides, anti-sense polynucleotides encoding them, kits, and their application to the in vitro detection, the in vivo and ex vivo of delivery of a therapeutic agent, the detection of the polynucleotides by hybridization with labeled probes, and the vaccination against and treatment of cancer and viral infections.
2. Description of the Background
The human milk fat globule (HMFG) has been used extensively as a source of antigenic material for the preparation of both polyclonal and monoclonal antibodies that have found widespread use in the diagnosis of breast cancer, as well as in the study of the breast epithelial cell surface and the processing of its antigenic components.
Polyclonal antiserum was originally prepared, that after appropriate absorptions with non-breast tissue was found to identify surface antigens of human mammary epithelial cells (HME-Ags). This antiserum (anti-HME) had a high specificity for normal breast epithelial cells and breast carcinomas. It identified mainly three components of the human milk fat globule which had molecular weights of 150 Kdalton, 70 Kdalton, and 46 Kdalton, respectively.
Monoclonal antibodies were first made against the HMFG in 1980. These antibodies were applied to identify a hitherto unknown component of the breast epithelial cell surface, a large molecular weight mucin-like glycoprotein, that was named non-penetrating glycoprotein (NPGP). This latter component appears to be extremely antigenic in the mouse. The vast majority of monoclonal antibodies prepared against HMFG as well as breast tumors have been found to have specificity against different epitopes of this mucin. Less frequently, monoclonal antibodies have been prepared against the 70 Kdalton and 46 Kdalton components of the HMFG.
The reason for the high immunogenicity of NPGP was elucidated by the characterization of cDNA clones selected from a .lambda.gt11 breast cell library using both polyclonal and monoclonal antibodies against the mucin. These cDNA clones consist of large arrays of highly conserved 60 bp tandem repeats. The resulting 20 amino acid repeat contains epitopes for several anti-mucin antibodies. The repeat is apparently unstable at the genomic level. This may account for the observed polymorphism seen at the gene, RNA and protein levels for this high molecular weight mucin. An initial report on cDNA cloning of the mucin product suggested that the core protein had a molecular weight of about 68 Kdalton. However, the mRNA was found to be large enough to code for proteins from about 170 Kdalton to 230 Kdalton. More recently, using milder deglycosylation methods, a core protein was identified having a molecular weight of about 200 Kdalton. Attention has also been devoted to the study and use of the NPGP mucin, largely as a result of its high immunogenicity. Thus, a large number of monoclonal antibodies were prepared against it. However, the smaller components of HMFG also appear to be important molecules on the surface of breast epithelial cells. They have a breast specificity as demonstrated by the anti-HMFG antibodies.
The 46 Kdalton and 70 Kdalton HMFG antigens are found in serum of breast cancer patients and thus can be used as markers for breast cancer in serum assays. In addition, the 70 Kdalton component has been found to co-purify with the intact mucin complex and has been reported to be associated with the NPGP mucin complex by means of disulfide bonds, making it a possible linker protein of this surface mucin complex. Polyclonal antibodies against a major component of the HMFG having molecular weight of 155 Kdaltons have been prepared. It was found that antisera bound also to the apical surface of lobules and terminal ducts, but not to the larger ducts of the mammary gland. The latter also did not bind to the apical surface of normal apocrine and eccrine sweat gland coils and ducts, or sebaceous glands in skin. The MFGM-gpl55 did become localized in Paget's disease and breast disease but not in cases of extramammary disease.
Few monoclonal antibodies, however, have been prepared against the smaller components of the HMFG system, such as the 70 Kdalton and 46 Kdalton HMFG antigens. The breast mucin glycoprotein molecule appears to be highly antigenic because of its internally repeated structure. The components of the mucin glycoprotein was recently determined and a partial sequence for the 70 Kdalton antigen obtained by cDNA cloning. A role for the 70 Kdalton antigen has been suggested as a linker protein for the breast mucin. The 46 Kdalton component of the HMFG system has been found to be present in the serum of breast cancer patients. In addition, with the aid of both monoclonal and polyclonal antibodies against the 46 Kdalton HMFG antigen, circulating immune complexes of the 46 Kdalton HMFG antigen were detected in breast cancer patients, and an increase in the circulating 46 Kdalton HMFG antigen was found to be associated with tumor burden.
Accordingly, there is still a need for an improved product and methods suitable for diagnostic and therapeutic applications to human cancer and virus-associated infections.